Large craniofacial bone defects remain a clinical challenge due to their complex shapes and large volumes. Stem cell-based technologies that deliver osteogenic stem cells have shown remarkable regenerative potential but are hampered by the need for extensive manipulation before implantation. To address this, we explored the bone forming potential of the clinically relevant stromal vascular fraction (SVF) cells obtained from human lipoaspirate. SVF cells can be isolated for acute use in the operating room and contain a subpopulation of adipose-derived stromal/stem cells (ASCs) that can develop mineralized tissue. ASCs can be purified from the more heterogeneous population of SVF cells via secondary and tertiary culture on tissue culture plastic. In this study, the relative potential for using SVF cells or passaged ASCs to induce robust bone regeneration was compared. Isogenic SVF and ASCs were suspended in fibrin hydrogels and seeded in three-dimensional-printed osteoinductive scaffolds of decellularized bone matrix and polycaprolactone. both cell populations successfully mineralized the scaffold, demonstrating the robust bone formation properties of SVF. In murine critical-sized cranial defects, ASC-loaded scaffolds had greater (but not statistically significant) bone volume and bone coverage area than SVF-loaded scaffolds. However, both cell-laden interventions resulted in significantly greater bone healing than contralateral acellular controls. In conclusion, we observed substantial mineralization and robust bone regeneration in tissue-engineered bone grafts using both SVF and passaged ASCs. Impact Statement The inability to effectively regenerate bone within critical-sized craniofacial defects is a present clinical challenge and overcoming this limitation using tissue engineering strategies would significantly advance current treatment outcomes. The present study tests the feasibility of harvesting stem cells intraoperatively, combining them with three-dimensional (3D)-printed osteoinductive scaffolds and, without culturing , implanting them into the bone defect to stimulate regeneration. The data from this study demonstrated that SVF isolated from lipoaspirate and used with minimal processing could be combined with a 3D-printed bioactive material in a point-of-care approach to promote bone regeneration.